We will investigate the fundamental parameters which control electron transfer reactions redox enzymes. Kinetic laser photolysis coupled with site-specific mutagenesis will elucidate the roles of amino acid side chains and other structural features in the generation of productive protein-protein or domain-domain interactions, and in electron transfer between prosthetic groups. Both interprotein and intraprotein reactions will be studied in the following specific systems. Ferredoxin/ferredoxin : NADP+ reductase: We will investigate the interactions of site-specific reductase mutants with both wild-type and mutant ferredoxins in order to provide new insights int the association/recognition and electron transfer events which occur during productive encounter Flavocytochrome b2 (lactate dehydrogenase): Conservative mutations will be made at the heme-flavin domain interface, and the effects of such changes on intramolecular electron transfer between the prosthetic groups will be determined. Coupled with x-ray structures of the mutant proteins, these studies should provide important insights into the role of distance and orientation on redox cents communication. Cytochrome P4SOBM-3 (fatty acid monooxygenase): The electron transfer properties of the holoenzyme will be compared with those of expressible subdomains of the protein (heme, FAD-FMN, FAD, FMN, and FMN-heme). This will provide information on the effects of interactions between these domains on intercenter electron transfer. We will also study the effect of mutations in the polypeptide linker region on interdomain electron transfer. p-Cresol methylhydroxylase ( flavocytochrome c): We will study intersubunit electron transfer kinetics in mutants which are alterated at the flavin site or in the subunit interface region. These studies, along with x-ray structure determinations of mutant proteins, should provide information on et pathways and distance and orientation effects on flavin-heme electron transfer.